Oxygen Sensing and Growth Plate Chrondrocyte Maturation

描述(申请人提供)：颅面复合体、头骨和下颌骨的生长依赖于软骨内生长板内软骨细胞的活动。生长板中细胞的成熟伴随着能量生成的特定阶段的变化。最近的研究表明，代谢控制是由缺氧诱导因子(HIF)介导的，HIF是一种对局部氧分压变化做出反应的转录因子。现已知道，HIF活性受一组作为氧感受器的脯氨酸羟基酶(PhDS)的控制。基于这些观察，我们推测在生长板中，PHD控制的HIF转录活性的激活是软骨细胞成熟、终末分化和诱导凋亡所必需的。这项建议中描述的研究旨在检验这一新的假说，该假说汇集了关于软骨细胞新陈代谢、局部环境的影响、终末分化状态的发展和细胞死亡机制的当前观点。因此，在特定的目标1中，我们测量了PHD在终末分化软骨细胞中的表达和分布，并确定了PHD表达的变化如何调节HIF的表达、HIF的转录活性和终末分化状态的发展。然后，我们将PHD的表达与小鼠骨盆生长板肥大状态的发展联系起来。在目标2中，我们确定了HIF-1a在终末分化软骨细胞中的表达和分布，并确定了HIF表达如何调节小鼠生长板中PHD亚型的表达。在目的3中，我们探讨了血氧状态的调节如何调控终末分化和诱导软骨细胞凋亡；我们在小鼠生长板中探讨了HIF-PhD的表达与软骨细胞凋亡/存活的关系。最后，在最后一个具体目标中，我们设计了有条件地过度表达PHD的转基因动物。我们使用这个模型来评估PHD表达对小鼠骨盆生长板发育的影响。我们认为HIF-1的过度表达与软骨细胞代谢和细胞凋亡/存活通路的活性有关。这项研究的结果应该有助于确定PHD和与软骨细胞成熟过程有关的下游事件之间的联系。

DESCRIPTION (provided by applicant): Growth of the craniofacial complex, the skull and the mandibular condyle is dependent on the activities of chondrocytes contained within the endochondral growth plate. Maturation of cells in the growth plate is accompanied by stage specific changes in energy generation. Recent studies show that metabolic control is mediated by Hypoxia Inducible Factor (HIF), a transcription factor that responds to changes in the local oxygen tension. It is now known that HIF activity is controlled by a group of prolyl hydroxylases (PHDs) which serve as oxygen sensors. Based on these observations, we hypothesize that in the growth plate, PHD controlled activation of HIF transcriptional activity is required for chondrocyte maturation, terminal differentiation and the induction of apoptosis. Investigations described in this proposal are aimed at testing this novel hypothesis that brings together current ideas concerning chondrocyte metabolism, the impact of the local environment, the development of the terminal differentiated state and the mechanism of cell death. Thus, in Specific Aim 1, we measure the expression and distribution of the PHDs in chondrocytes undergoing terminal differentiation and ascertain how changes in PHD expression modulate HIF expression, HIF transcriptional activity and development of the terminally differentiated state. We then relate PHD expression to development of the hypertrophic state in the mouse epiphyseal growth plate. In Aim 2, we determine the expression and distribution of HIF-la in chondrocytes undergoing terminal differentiation and ascertain how HIF expression modulates the expression of each of the PHD isoforms in the mouse growth plate. In Aim 3, we explore how modulation of the oxemic status regulates terminal differentiation and the induction of chondrocyte apoptosis; we relate HIF-PHD expression to chondrocyte apoptosis/survival in the mouse growth plate. Finally, in the last Specific Aim, we engineer transgenic animals that conditionally over-express the PHDs. We use this model to assess the impact of PHD expression on the development of the murine epiphyseal growth plate. We relate over-expression to HIF-1 expression, chondrocyte metabolism and the activity of the apoptosis/survival pathways. Outcomes from this investigation should help define the linkage between PHDs and downstream events linked to the chondrocyte maturation process.